Recently, the existence of surface active substance in the eustachian tube has been reported. The chemical composition and surface tension lowering properties of surface tension lowering substance(STLS) were found to be almost identical in both lung and eustachian tube. However, specific secretory site of eustachian STLS have not so far been studied extensively. Authors investigated secretory activities and sites of the tubotympanum lining of normal conditions in the rabbit by using autoradiography with 3H-palmitic acid on the eustachian tube. This investigation is a part of the primary study to establish the dynamics of cell secretion. The epithelium of eustachian tube is covered with ciliated cell, nonciliated cell, basal cell and secretory cell. A large amount of isotope was observed in the secretory cells within the epithelium of eustachian tube. The result suggests that the secretory cell is the main site for producing eustachian tube STLS. A further investigation for the secretory site of eustachian tube STLS by using electron microscopic autoradiography will be necessary.